The present invention relates to a method and an apparatus for spectrophotometrically determining the concentration of a number of hemoglobin derivatives (or parameters derived from the concentration of individual hemoglobin derivatives) in whole blood.
It is known to determine the concentration of various hemoglobin derivatives such as deoxyhemoglobin, oxyhemoglobin, carboxyhemoglobin, methemoglobin, and sulfhemoglobin by spectrophotometric methods (cf. e.g., Ole Siggaard-Andersen, The Acid-Base Status of the Blood, 4th edition, Munksgaard, Copenhagen, 1974, pp. 181-187, and U.S. Pat. No. 3,927,614). Information concerning the concentration of the various hemoglobin derivatives, in particular oxyhemoglobin and deoxyhemoglobin, is important in addition to information on the oxygen concentration or oxygen partial pressure of the blood in several clinical situations such as, e.g., when a patient is under anaesthesia.
As explained in Siggaard-Andersen, loc.cit., the theoretical basis for the spectrophotometric determination of the concentration of hemoglobin derivatives in a mixture is that the total absorbance at any wavelength is the sum of the contribution of each derivative, and that each derivative obeys Lambert-Beer's law. Thus, ##EQU1## wherein A.sup..lambda. is the absorbance of the mixture at the wavelength .lambda., C.sub.y is the concentration of derivative y, l is the light path length, and .epsilon..sub.y.sup..lambda. is the extinction coefficient of derivative y at wavelength .lambda..
The absorbance A is defined as ##EQU2## wherein lo is the intensity of the incident light on the sample, and l is the intensity of the resultant modified light. The ratio l/lo is designated the transmittance.
In accordance with this, the concentrations of a certain number of hemoglobin derivatives in a blood sample may be determined by measuring the transmittance or the absorbance of the blood sample at at least the same number of different wavelengths. Provided that the path length of the light and data representative of the extinction coefficient for each derivative at each wavelength are known, the unknown concentrations may be determined by solving an equation set of the certain number of linear equations with the same number of variables.
While the known methods for determining hemoglobin derivatives (e.g. the methods described in U.S. Pat. No. 3,972,614) function satisfactorily on most blood samples, wrong results will be obtained when using these methods in connection with turbidity-containing blood samples. Thus, Siggaard-Andersen, loc.cit., pp. 186-187, mentions that turbidity of the hemolyzate, caused by leukocytes, lipemia, or erythrocyte ghosts, is a major source of error. A recent article, "Effect of Intralipid on Measurements of Total Hemoglobin and Oxyhemoglobin in Whole Blood" Lakshman R. Sehgal et al., Critical Care Medicine, Vol. 12, 10, October 1984, pp. 907-909 deals with the problems incurred by the light scattering effect on the determination of hemoglobin species on blood from patients who have received Intralipid.RTM. (an infusion liquid containing, inter alia, soy lecithin and glycerol). The article states that the light scattering affects the measured concentrations of all hemoglobin species, but most dramatically the methemoglobin, and concludes that the data for hemoglobin derivatives measured on blood samples from patients who have received Intralipid.RTM. should be disregarded.
Thus, the presence of turbidity in blood subjected to hemoglobin derivative determination presents a major problem which has, until now, not been solved in a satisfactory manner.
It has now been found that the error derived from the existence of turbidity in whole blood under test can be eliminated to a completely satisfactory degree, thereby making it possible to obtain reliable data for the concentration of the various hemoglobin derivatives irrespective of whether the blood in question contains turbidity-incurring components or not.